Objective: To downregulate the expression of mdr1 gene in K562/A02 cell line by RNA interference.
Methods: The eukaryotic expression vectors of shRNA aiming at two mdr1 mRNA target sequences were constructed and used to transfect the drug resistance cell line K562/A02 with liposome-induced gene transfection. The mRNA of mdr1 gene was identified by Q-PCR. The P-gp expression of was detected by Western blot. The function of P-gp was measured by daunorubicin (DNR) efflux experiment and the sensitivity of cell lines to doxorubicin (ADM) was detected by MTT test.
Results: Two shRNA plasmids targeting mdr1 mRNA were constructed and cloned. Two mdrl-targeted shRNA could down-regulate mdr1 mRNA expressions with 89.74% and 87.18% respectively, almost completely down-regulate the P-gp expression. The intracellular DNR increased after RNAi treating. The daunorubicin efflux ratio at 60 min were 13.16% and 22.02%, compared with control 40.44% , 45.31%, P<0.05. MTT test demonstrated the relative reversing efficiencies to doxorubicin were 84.36% and 76.69% respectively. CONCLUSION RNA interference can effectively reverse multidrug resistance caused by mdr1.